Research Group Nanotechnology for Life Science, Fraunhofer Institute for Applied Polymer Research, Geiselbergstrasse 69, Potsdam-Golm 14476, Germany.
Biomacromolecules. 2010 Aug 9;11(8):2130-5. doi: 10.1021/bm1005036.
Thermoresponsive oligo(ethylene glycol)-based copolymers were investigated for trypsin conjugation. These copolymers have been synthesized by atom transfer radical polymerization of 2-(2-methoxyethoxy)ethyl methacrylate (MEO(2)MA) with oligo(ethylene glycol) methyl ether methacrylate (OEGMA(475), M(n) = 475 g.mol(-1)) at 60 degrees C in the presence of copper(I) chloride and 2,2'-bipyridyl. Two different ATRP initiators, containing succinimidyl ester moieties, were tested, namely, N-succinimidyl-2-bromopropionate and N-succinimidyl-2-bromoisobutyrate. In both cases, ATRP afforded well-defined polymers with a narrow molecular weight distribution and controlled chain-ends. However, the efficiency of initiation of the two initiators was lower than 1 and therefore the formed polymers exhibited a higher than expected mean degree of polymerization. Nevertheless, all types of polymers could be conjugated to trypsin. The conjugation reaction was performed in borax-HCl buffer. Sodium dodecyl sulfate poly(acrylamide) gel electrophoresis (SDS-PAGE) indicated that polymer/enzyme conjugates were obtained in all cases. However, (co)polymers initiated by N-succinimidyl-2-bromopropionate led to the best conjugation results. The formed P(MEO(2)MA-co-OEGMA(475))-trypsin conjugates were found to be thermoresponsive and moreover exhibited a higher enzymatic activity than unmodified trypsin.
研究了温敏性聚乙二醇基共聚物用于胰蛋白酶的偶联。这些共聚物是通过在铜（I）氯化物和 2,2'-联吡啶存在下，在 60°C 下用 2-（2-甲氧基乙氧基）乙基甲基丙烯酸酯（MEO（2）MA）与聚乙二醇甲基醚甲基丙烯酸酯（OEGMA（475），M（n）= 475 g.mol（-1））进行原子转移自由基聚合合成的。测试了两种不同的 ATRP 引发剂，均含有琥珀酰亚胺酯部分，即 N-琥珀酰亚胺基-2-溴代丙酸酯和 N-琥珀酰亚胺基-2-溴代异丁酸酯。在这两种情况下，ATRP 都得到了具有窄分子量分布和受控链端的良好定义的聚合物。然而，两种引发剂的引发效率均低于 1，因此形成的聚合物的平均聚合度高于预期。尽管如此，所有类型的聚合物都可以与胰蛋白酶偶联。偶联反应是在硼砂-HCl 缓冲液中进行的。十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）表明，在所有情况下都获得了聚合物/酶缀合物。然而，由 N-琥珀酰亚胺基-2-溴代丙酸酯引发的（共）聚合物导致了最佳的偶联结果。形成的 P（MEO（2）MA-co-OEGMA（475））-胰蛋白酶缀合物表现出温敏性，并且比未修饰的胰蛋白酶具有更高的酶活性。
